The Thioesterase Bhp is Involved in the Formation of β‐Hydroxytyrosine during Balhimycin Biosynthesis in Amycolatopsis balhimycina
2010
The putative hydrolase gene bhp from the balhimycin biosynthetic gene cluster has been cloned and overexpressed
in Escherichia coli. The corresponding enzyme Bhp was purified to homogeneity by nickel-chelating
chromatography and characterized. Although Bhp has sequence similarities to hydrolases with
“haloperoxidase”/perhydrolase activity, it did not show any enzymatic activity with standard
“haloperoxidase”/perhydrolase substrates (e.g., monochlorodimedone and phenol red), nonspecific esterase
substrates (such as p-nitrophenyl acetate, p-nitrophenyl phosphate and S-thiophenyl acetate) or the model
lactonase substrate dihydrocoumarin. However, Bhp could be shown to catalyse the hydrolysis of
S-β-hydroxytyrosyl-N-acetyl cysteamine thioester (β-OH-Tyr-SNAC) with 15 times the efficiency of S-L-tyrosyl-
N-acetyl cysteamine thioester (L-Tyr-SNAC). This is in agreement with the suggestion that Bhp is involved in
balhimycin biosynthesis, during which it was supposed to catalyse the hydrolysis of β-OH-Tyr-S-PCP
(PCP=peptidyl carrier protein) to free β-hydroxytyrosine (β-OH-Tyr) and strongly suggests that Bhp is a
thioesterase with high substrate specificity for PCP-bound β-OH-Tyr and not a “haloperoxidase”/perhydrolase or
nonspecific esterase.
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